Sodium ion detection method

Abstract

The invention discloses a sodium ion detection method. The sodium ion detection method comprises the following steps: S1, testing the independent absorbance of one of two reagents; s2, mixing the two reagents, testing the mixed absorbance after mixing the two reagents, and obtaining the absorbance change rate of the reagents; s3, adding the sample into the mixed reagent, respectively testing the initial absorbance and the final absorbance, and obtaining the reaction absorbance change rate after the sample is added; s4, subtracting the absorbance change rate of the reagent obtained in the step S2 from the absorbance change rate of the reaction obtained in the step S3 to obtain the final absorbance change rate; s5, substituting the final absorbance change rate into the standard curve to obtain the sodium ion concentration of the to-be-detected sample. According to the scheme provided by the invention, the concentration value of sodium ions in the sample can be accurately detected.

Description

Technical Field

[0001] The present application relates to the technical field of detection methods, and in particular to a method for detecting sodium ions. Background Art

[0002] A serum sodium test is a routine test that assesses electrolyte balance by measuring the concentration of sodium ions (Na⁺) in the blood. It is primarily used to assist in the diagnosis of dehydration, edema, kidney disease, or endocrine abnormalities. Sodium is a key electrolyte in maintaining fluid distribution, neuromuscular function, and blood pressure. Abnormalities in sodium may indicate a variety of diseases or metabolic disorders.

[0003] Common sodium ion detection methods include flame photometry, ion-selective electrode (ISE), and biochemical methods. The biochemical method works as follows: sodium-dependent β-galactosidase catalyzes the substrate o-nitrophenol-β-D-galactoside (ONPG) to produce o-nitrophenol and galactose. The increase in the absorbance of o-nitrophenol at 405 nm is proportional to the sodium ion concentration, and the sodium ion concentration is determined by the change in absorbance. The biochemical method can be performed using existing spectrophotometers or automated biochemical analyzers. It is low-cost and fast-response, making it particularly suitable for meeting the large-volume sample testing needs of hospitals, testing centers, and other institutions.

[0004] However, there are still certain problems with the stability of the biochemical method, which can easily lead to deviations in the test results. Summary of the Invention

[0005] In order to solve the above technical problems, the purpose of the present invention is to provide a method for detecting sodium ions. The solution provided in this application is to maintain the final signal value within a deviation range that meets the requirements by subtracting the signal value change rate of the test reagent itself from the signal value change rate before and after the addition of the sample to react; that is, the concentration value of sodium ions in the sample can be accurately detected.

[0006] The technical solutions provided by the present invention are as follows: A method for detecting sodium ions comprises the following steps: S1. Test the absorbance of one of the two reagents alone; S2. Mix the two reagents, test the mixed absorbance of the two reagents, and obtain the absorbance change rate of the reagents; S3. Add the sample to the mixed reagent, measure the initial absorbance and the final absorbance, and obtain the rate of change of the reaction absorbance after adding the sample; S4, subtracting the absorbance change rate of the reagent obtained in step S2 from the absorbance change rate of the reaction obtained in step S3 to obtain the final absorbance change rate; S5. Substitute the final absorbance change rate into the standard curve to obtain the sodium ion concentration of the sample to be tested.

[0007] Preferably, in step S2, after the two reagents are mixed, wait for the first period of time, and then measure the mixed absorbance; In step S3, after the sample is added to the mixed reagent, wait for the second period of time, measure the initial absorbance, and then wait for the third period of time to measure the final absorbance.

[0008] Preferably, the first period of time is 1 - 3 min, the second period of time is 1 - 3 min, and the third period of time is 3 - 5 min.

[0009] Preferably, the first period of time is 1 min, the second period of time is 1 min, and the third period of time is 3 min.

[0010] Preferably, in step S5, the standard curve is established according to the following method: Use multiple standard samples with different concentrations, and perform tests according to the methods of steps S1 to S4 to obtain the reaction absorbance change rate corresponding to each concentration of the standard sample. Establish a standard curve based on the concentrations of multiple standard samples and their corresponding reaction absorbance change rates.

[0011] Preferably, establishing a standard curve based on the concentrations of multiple standard samples and their corresponding reaction absorbance change rates specifically includes: Using multiple reaction absorbance change rates as the abscissa and different concentrations of multiple standard samples as the ordinate, perform linear fitting to obtain the standard curve.

[0012] Preferably, reagent R1 includes β - galactosidase, buffer, stabilizer, and cryptand; Reagent R2 includes o - nitrophenyl - β - D - galactoside, buffer, and stabilizer.

[0013] Preferably, the buffer is any one of tris - hydroxymethyl aminomethane, 3 - morpholinopropanesulfonic acid, and 4 - (2 - hydroxyethyl)piperazine - 1 - ethanesulfonic acid; The stabilizer is any one of ethylene glycol, glycerol, bovine serum albumin, mannitol, trehalose, and sucrose; The cryptand is any one of 18 - crown - 6, 15 - crown - 5, and 12 - crown - 4.

[0014] Preferably, reagent R1 includes β - galactosidase at a concentration of 0.5 - 5 KU / L, buffer at a concentration of 10 - 150 mM, stabilizer at a concentration of 0.5 - 10 g / L, and cryptand at a concentration of 2 - 10 mM; Reagent R2 includes o - nitrophenyl - β - D - galactoside at a concentration of 2 - 10 mM, buffer at a concentration of 10 - 150 mM, and stabilizer at a concentration of 0.5 - 10 g / L.

[0015] Preferably, the absorbance is measured at the following wavelengths: main wavelength 405 nm, secondary wavelength 660 nm; The reaction temperature is 37 - 40 °C.

[0016] In the research, the applicant found that when using the biochemical method to detect the sodium ion concentration, the deviation occurred because the stability of the reagent used in the detection decreased after long-term storage. Specifically, the galactosidase in the detection reagent was unstable. When using the rate method for testing, the decrease in its enzyme activity would cause the sodium ion concentration to be lower than the true value. However, if the reagent was re-prepared every time for detection, the operation was troublesome and the implementation was difficult.

[0017] The applicant found that by changing the reagent addition order and finding the relationship between the change rate of the reagent absorbance and the change rate of the absorbance before and after the sample addition reacted, the problem of low test values caused by the decrease in reagent stability could be improved. Specifically, in the solution provided in this application, by subtracting the change rate of the signal value of the test reagent itself from the change rate of the signal value before and after the sample addition reacted, the finally obtained signal value was maintained within a range where the deviation met the requirements; that is, the concentration value of sodium ions in the sample could be accurately detected.

[0018] Specifically, the operation according to the method provided in this application is as follows: S1. Test the individual absorbance of one of the two reagents; S2. Mix the two reagents and test the mixed absorbance of the two reagents after mixing to obtain the change rate of the reagent absorbance; S3. Add the sample to the mixed reagent and test the initial absorbance and the final absorbance respectively to obtain the change rate of the reaction absorbance after adding the sample; S4. Subtract the change rate of the reagent absorbance obtained in step S2 from the change rate of the reaction absorbance obtained in S3 to obtain the final change rate of the absorbance; S5. Substitute the final change rate of the absorbance into the standard curve to obtain the sodium ion concentration of the sample to be tested.

[0019] Preferably, in step S2, after the two reagents are mixed, wait for the first time and then test the mixed absorbance; in step S3, after the sample is added to the mixed reagent, wait for the second time, test the initial absorbance, and then wait for the third time to test the final absorbance.

[0020] The change rate of the absorbance is a well-known concept in the art. Specifically, the difference between the mixed absorbance and the individual absorbance, divided by the first time, is the change rate of the reagent absorbance. The difference between the final absorbance and the initial absorbance, divided by the third time, is the change rate of the reaction absorbance.

[0021] Representing each parameter with symbols, the method for detecting sodium ions provided in this application is as follows: S1. Test the individual absorbance A0 of reagent R1 or reagent R2. S2. Mix reagent R2 with reagent R1 in step S1, or mix reagent R1 with reagent R2 in step S1, wait for the first time T1, and then detect the absorbance A1 of the mixed reagent. Calculate the change rate C1 of the reagent absorbance, where C1 = (A1 - A0) / T1; S3. Add the sample to the mixed reagent, wait for the second time T2, measure the absorbance B0, then wait for the third time T3, and measure the absorbance B1; Calculate the change rate C2 of the reaction absorbance, where C2 = (B1 - B0) / T3; S4. Calculate the final change rate C3 of the absorbance, where C3 = C2 - C1; S5. Substitute C3 into the standard curve to obtain the sodium ion concentration of the sample to be measured.

[0022] In this application, the change rate C1 of the reagent absorbance is (A1 - A0) / T1, and the change rate C2 of the reaction absorbance is (B1 - B0) / T3, which reflects the change in absorbance per unit time and is not affected by the specific time lengths of T1 and T3 during the detection process. As long as the reaction time meets the conventional time-consuming range for sodium ion detection, it is fine.

[0023] In this application, whether it is to detect the individual absorbance A0 of reagent R1 or the individual absorbance A0 of reagent R2 in step S1, after that, it is necessary to perform operations with the absorbance A1 of the mixed reagent to eliminate the influence brought by the change in reagent components. Therefore, whether the individual absorbance A0 comes from reagent R1 or reagent R2 has no influence on the accuracy of the test results.

[0024] The method for detecting sodium ions provided in this application only changes the use of the absorbance signal and does not make any changes to the reagents and samples, and is applicable to the detection using sodium ion detection reagents well-known in the art. Specific parameters such as the formula and concentration of the sodium ion reagent do not constitute a limitation to the solution of this application. Detailed implementation mode

[0025] In order to enable those skilled in the art to better understand the technical solutions in this application, the technical solutions in the embodiments of this application will be clearly and completely described below. Obviously, the described embodiments are only a part of the embodiments of this application, rather than all the embodiments. Based on the embodiments in this application, all other embodiments obtained by those of ordinary skill in the art without creative efforts shall fall within the scope of protection of this application.

[0026] The reagents used in the examples and comparative examples are shown in the following table:

[0027]

[0028] The volume ratios of reagent R1, reagent R2 to the sample are 200:80:8 (unit: μl).

[0029] The detection process was carried out on a fully automatic biochemical analyzer, and the parameters were set as follows: reaction temperature 37°C, main wavelength 405 nm, and secondary wavelength 660 nm. The detection methods described in Example 1 and Comparative Example 1 were respectively used for detection.

[0030] Example 1 A method for detecting sodium ions includes the following steps: S1. Measure the absorbance A0 of the test reagent R1 alone; S2. Mix reagent R2 with reagent R1 in step S1, wait for the first time T1 (1 min), and then measure the absorbance A1 of the mixed reagent. Calculate the reagent absorbance change rate C1, C1 = (A1 - A0) / T1 = (A1 - A0); S3. Add the sample to the mixed reagent, wait for the second time T2 (1 min), measure the absorbance B0, and then wait for the third time T3 (3 min), and measure the absorbance B1; Calculate the reaction absorbance change rate C2, C2 = (B1 - B0) / T3 = (B1 - B0) / 3; S4. Calculate the final absorbance change rate C3, where C3 = C2 - C1; S5. Substitute C3 into the standard curve to obtain the sodium ion concentration of the sample to be tested.

[0031] Among them, in step S5, the standard curve was established according to the following method: Use standard samples and test according to the methods of steps S1 to S4 to establish the standard curve.

[0032] Comparative Example 1 S1. Mix the sample with reagent R1 and wait for 5 min; S2. Add reagent R2, record the absorbance A1' after 1 min, and record the absorbance A2' after reacting for 3 min; S3. Calculate the absorbance change rate C1 between two points, C1 = (A2' - A1') / 3; S4. Detect the corresponding reaction absorbance change rate B2' of the sample to be tested according to the methods of steps S1 to S3, and substitute it into the standard curve to obtain the sodium ion concentration of the sample to be tested.

[0033] Among them, in step S4, the standard curve was established according to the following method: Use multiple standard samples with different concentrations and test according to the methods of steps S1 to S3 to establish the standard curve.

[0034] According to the methods of Example 1 and Comparative Example 1, tests were conducted using freshly prepared reagents (denoted as 0 months), as well as reagents stored for 2, 4, 6, 8, 10, and 12 months, and the deviation from the test value at the 0th month was calculated. The results are shown in Table 1-2.

[0035] Table 1 Detection Results of the Method of Comparative Example 1

[0036] During the applicant's research on existing methods, for reagents stored for different times, absorbance detection of the reagents themselves was also carried out to explore the factors affecting absorbance changes. The detection results of the individual absorbance A0 of reagent R1 are shown in Table 2: Table 2

[0037] From the changes in absorbance measured at different times in Table 1 and Table 2, it can be seen that for the conventional test method of Comparative Example 1, the ratio of the change amplitudes of reagent R1 and absorbance change rate C1 is close to 1:1. Therefore, the applicant determined that the decrease in absorbance caused by the decrease in β-galactosidase in reagent component R1 is strongly correlated with the decrease in the reagent reaction rate. According to the conventional scheme of Comparative Example 1 for testing, after 12 months of storage, the test value of the reagent is about 16% lower than that at the 0th month.

[0038] Table 3 Detection Results of the Method of Example 1

[0039] The applicant adjusted the reagent addition order, the light signal acquisition order, and optimized the absorbance calculation method to obtain the test method of Example 1. It can be seen from Table 3 that the decrease amplitude of the reagent system (reagent 1 + sample + reagent 2) represented by the reaction absorbance change rate C2 is close to the decrease amplitude of the mixture of reagent 1 and reagent 2 represented by the reagent absorbance change rate C1 during this process. Therefore, the signal value obtained by subtracting the two (i.e., the final absorbance change rate C3) is less affected by reagent stability, optimizing the sodium ion test method and the calculation method of the signal value, and greatly improving the impact of the decrease within the reagent shelf life on the test value.

[0040] The above description of the disclosed embodiments enables those skilled in the art to implement or use the present invention. Various modifications to these embodiments will be obvious to those skilled in the art, and the general principles defined herein can be implemented in other embodiments without departing from the spirit or scope of the present invention. Therefore, the present invention will not be limited to these embodiments shown herein, but rather will be accorded the widest scope consistent with the principles and novel features disclosed herein.

Claims

1. A method for detecting sodium ions, characterized in that, It includes the following steps: S1. Measure the absorbance of one of the two reagents alone; S2. Mix the two reagents and measure the absorbance of the mixture after mixing to obtain the reagent absorbance change rate; S3. Add the sample to the mixed reagent, measure the initial absorbance and the final absorbance respectively to obtain the reaction absorbance change rate after adding the sample; S4. Subtract the reagent absorbance change rate obtained in step S2 from the reaction absorbance change rate obtained in S3 to obtain the final absorbance change rate; S5. Substitute the final absorbance change rate into the standard curve to obtain the sodium ion concentration of the sample to be tested.

2. The sodium ion detection method according to claim 1, characterized in that In step S2, after the two reagents are mixed, wait for the first period of time and then measure the absorbance of the mixture; In step S3, after the sample is added to the mixed reagent, wait for the second period of time, measure the initial absorbance, and then wait for the third period of time to measure the final absorbance.

3. The sodium ion detection method according to claim 2, wherein The first period of time is 1 - 3 min, the second period of time is 1 - 3 min, and the third period of time is 3 - 5 min.

4. The method for detecting sodium ions according to claim 3, wherein The first period of time is 1 min, the second period of time is 1 min, and the third period of time is 3 min.

5. The method for detecting sodium ions according to any one of claims 1-4, characterized in that, In step S5, the standard curve is established according to the following method: Use multiple standard samples with different concentrations, test them according to the methods of steps S1 to S4 to obtain the reaction absorbance change rate corresponding to each concentration of the standard sample, and establish a standard curve based on the concentrations of multiple standard samples and their corresponding reaction absorbance change rates.

6. The sodium ion detection method according to claim 5, characterized in that Establish a standard curve based on the concentrations of multiple standard samples and their corresponding reaction absorbance change rates, specifically including: Use multiple reaction absorbance change rates as the abscissa and the different concentrations of multiple standard samples as the ordinate for linear fitting to obtain the standard curve.

7. The sodium ion detection method according to claim 1, characterized in that, Reagent R1 includes β-galactosidase, buffer, stabilizer and cryptand; Reagent R2 includes o-nitrophenyl-β-D-galactoside, buffer and stabilizer.

8. The method for detecting sodium ions according to claim 7, characterized in that, The buffer is any one of tris(hydroxymethyl)aminomethane, 3-(N-morpholino)propanesulfonic acid, 4-(2-hydroxyethyl)-1-piperazineethanesulfonic acid; The stabilizer is any one of ethylene glycol, glycerol, bovine serum albumin, mannitol, trehalose, sucrose; The cryptand is any one of 18-crown-6, 15-crown-5, 12-crown-4.

9. The method for detecting sodium ions according to claim 8, wherein, Reagent R1 includes β-galactosidase at a concentration of 0.5 - 5 KU / L, buffer at a concentration of 10 - 150 mM, stabilizer at a concentration of 0.5 - 10 g / L and cryptand at a concentration of 2 - 10 mM; 10. The sodium ion detection method according to claim 1, wherein, Reagent R2 includes o-nitrophenyl-β-D-galactoside at a concentration of 2 - 10 mM, buffer at a concentration of 10 - 150 mM and stabilizer at a concentration of 0.5 - 10 g / L. The absorbance is measured at the following wavelengths: main wavelength 405 nm, secondary wavelength 660 nm; The reaction temperature is 37 - 40 °C.

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